Levothyroxine liquid formulations

ABSTRACT

The present invention is directed to a liquid formulation comprising levothyroxine or a pharmaceutically acceptable salt thereof. The formulation of the present invention includes tromethamine, sodium iodide, and water and has a pH of about 9.0 to about 11.5. The liquid formulation according to the invention is stable and ready-to-use.

CROSS-REFERENCE TO RELATED APPLICATIONS

This patent application is a continuation of co-pending U.S. patentapplication Ser. No. 15/366,864, filed Dec. 1, 2016, the disclosure ofwhich is incorporated herein by reference in its entirety for allpurposes.

BACKGROUND OF THE INVENTION

Levothyroxine sodium for injection is a sterile lyophilized product forparenteral administration of levothyroxine sodium for thyroidreplacement therapy. Levothyroxine sodium for injection is particularlyuseful when thyroid replacement is needed on an urgent basis, for shortterm thyroid replacement, and/or when oral administration is notpossible, such as for a patient in a state of myxedema coma.

Full chemical names for levothyroxine sodium include4-(4-hydroxy-3,5-diiodophenoxy)-3,5-diiodo-L-phenylalanine sodium, andL-tyrosine-O-(4-hydroxy-3,5-diiodophenyl)-3,5-diiodo-monosodium salt.Levothyroxine sodium has a molecular weight of approximately 798.85 andthe following chemical structure:

Conventional formulations of levothyroxine sodium for injection arepreservative-free lyophilized powders containing levothyroxine sodiumand the excipients mannitol, sodium phosphate buffer, and sodiumhydroxide. Administration of the conventional formulations involvereconstitution of the lyophilized powder in 0.9% sodium chlorideinjection (USP) to provide an injectable solution.

However, use of the conventional lyophilized formulations requiresreconstitution or dilution by healthcare practitioners prior to use.Once reconstituted, the levothyroxine sodium solutions have a limitedstability, and must be used within a few hours of reconstitution. Inaddition, contaminants may be introduced into the solutions during thereconstitution process, thereby compromising patient safety.

It has been shown that levothyroxine in oral tablets and in aqueoussolutions undergoes degradation. Major degradation products oflevothyroxine are known to include 3,3′,5-triiodothyronine (T3)3,5-diiodothyronine (T2) 3,3′,5,5′-tetraiodothyroacetic acid (TTAA4)3,3′,5-triiodothyroacetic acid (TTAA3) and 3,5-diiodothyroacetic acid(TTAA2) (Kannamkumarath et al., J. Anal. At. Spectrom., 2004, 19:107-113 and Patel et al., Int. J. Pharm., 2003, 264: 35-43)).3,3′,5-triiodothyronine, known as liothyronine or T3, is a majordegradant. Aqueous solutions of levothyroxine sodium have been shown tobe more stable at basic pH than at acidic pH, but significantdegradation of levothyroxine sodium also has been shown to occur atbasic pH (Patel et al., Int. J. Pharm., 2003, 264: 35-43).

Thus, there remains a need in the art for a ready-to-use injectableformulation of levothyroxine sodium that exhibits storage stability.

BRIEF SUMMARY OF THE INVENTION

The invention provides a liquid formulation comprising levothyroxine ora pharmaceutically acceptable salt thereof, tromethamine, sodium iodide,and water, wherein the formulation has a pH of about 9.0 to about 11.5.

The invention also provides a liquid formulation comprising (a)levothyroxine or a pharmaceutically acceptable salt thereof in aconcentration of about 20 mcg/mL to about 100 mcg/mL, (b) tromethaminein a concentration of about 5 mg/mL to about 20 mg/mL, (c) sodium iodidein a concentration of about 100 mcg/mL to about 300 mcg/mL, (d) sodiumchloride, and (e) water, wherein the formulation has a pH of about 9.8to about 10.8.

DETAILED DESCRIPTION OF THE INVENTION

The invention provides a liquid formulation comprising levothyroxine ora pharmaceutically acceptable salt thereof, tromethamine, sodium iodide,and water, wherein the formulation has a pH of about 9.0 to about 11.5.The liquid formulation according to the invention is stable andready-to-use.

As used herein, a “ready-to-use” formulation is a sterile, injectableformulation that is not reconstituted from a solid by a healthcareprovider prior to use. Rather, a ready-to-use formulation is supplied bya pharmaceutical manufacturer in a suitable container (e.g., vial,syringe, bag, container) in liquid form. In some embodiments, aready-to-use formulation is an injectable formulation that isadministered to a subject without dilution. In other embodiments, aready-to-use formulation is a concentrated, liquid solution that must bediluted prior to administration to a subject. Thus, in some embodiments,the formulation of the present invention can be further diluted in anappropriate diluent such as, for example, WFI (water for injection),0.9% sodium chloride, or 5% dextrose to a lower levothyroxineconcentration.

The formulation according to the present invention is stable. As usedherein, the terms “stable” and “stability” encompass any characteristicof the formulation which may be affected by storage conditionsincluding, without limitation, potency, total impurities, levothyroxinedegradation products, specific optical rotation, optical purity, watercontent, appearance, viscosity, sterility, and color and clarity. Thestorage conditions which may affect stability include, for example,duration of storage, temperature, humidity, and/or light exposure.

In certain embodiments, a stable levothyroxine formulation refers to aformulation that retains at least about 90%, or about least about 95%,or at least about 96%, or at least about 98%, of the labeledconcentration of levothyroxine or pharmaceutically acceptable saltthereof after storage under typical and/or accelerated conditions. Infurther embodiments, a stable levothyroxine formulation refers to lessthan about 15% (area percent), or less than about 10% (area percent), orless than about 7% (area percent), or less than about 5% (area percent),or less than about 2% (area percent) of levothyroxine-related impuritiesare present after storage under typical and/or accelerated conditions.

In some embodiments, the liquid formulation of the invention is stablefor at least 12 months, at least 18 months, at least 24 months, or atleast 36 months at refrigerated temperature (e.g., at 5±2° C.). In otherembodiments, the liquid formulation of the invention is stable for atleast 12 months, at least 18 months, at least 24 months, or at least 36months at room temperature (e.g., at 25±2° C.).

Methods for determining the stability of a formulation of the inventionwith respect to a given parameter are well-known to those of skill inthe art. For example, individual impurities and total impurities can beassessed by high-performance liquid chromatography (HPLC) or thin layerchromatography (TLC). Unless otherwise indicated to the contrary, apercentage amount of liothyronine, other individual impurities, or totalimpurities reported herein in the formulation is determined by a peakarea percent method using HPLC.

The formulation comprises levothyroxine or any pharmaceuticallyacceptable salt thereof. Preferably, the formulation compriseslevothyroxine sodium. In an embodiment, the levothyroxine sodium islevothyroxine sodium pentahydrate, which is the sodium salt of thelevo-isomer of thyroxine, an active physiological substance found in thethyroid gland.

When the formulation comprises levothyroxine sodium, the levothyroxinesodium can be present in the formulation in any suitable concentration.Typically, levothyroxine sodium can be present in the formulation at aconcentration of about 5 mcg/mL (micrograms/milliliter) or more, forexample, about 10 mcg/mL or more, about 15 mcg/mL or more, about 20mcg/mL or more, about 25 mcg/mL or more, about 30 mcg/mL or more, about35 mcg/mL or more, about 40 mcg/mL or more, or about 45 mcg/mL or more.

Alternatively, levothyroxine sodium can be present in the formulation ata concentration of about 500 mcg/mL or less, for example, about 450mcg/mL or less, about 400 mcg/mL or less, about 350 mcg/mL or less,about 300 mcg/mL or less, about 250 mcg/mL or less, about 200 mcg/mL orless, or about 150 mcg/mL or less.

Levothyroxine sodium can be present in the formulation in aconcentration bounded by any two of the aforementioned endpoints. Forexample, levothyroxine sodium can be present in the formulation in aconcentration of about 5 mcg/mL to about 500 mcg/mL, for example, about10 mcg/mL to about 450 mcg/mL, about 15 mcg/mL to about 400 mcg/mL,about 20 mcg/mL to about 350 mcg/mL, about 25 mcg/mL to about 300mcg/mL, about 30 mcg/mL to about 300 mcg/mL, about 35 mcg/mL to about300 mcg/mL, about 40 mcg/mL to about 300 mcg/mL, about 45 mcg/mL toabout 300 mcg/mL, or about 50 mcg/mL to about 250 mcg/mL, or about 20mcg/mL to about 100 mcg/mL.

In a preferred embodiment, levothyroxine sodium is present at aconcentration of about 20 mcg/mL. In another preferred embodiment,levothyroxine sodium is present at a concentration of about 40 mcg/mL.In yet another preferred embodiment, levothyroxine sodium is present ata concentration of about 100 mcg/mL.

The formulation can be provided in any suitable volume. In someembodiments, the volume of the formulation is about 0.5 mL or more,e.g., about 1 mL or more, about 3 mL or more, about 5 mL or more, about8 mL or more, about 10 mL or more, about 20 mL or more, or about 50 mLor more. In other embodiments, the volume of the formulation is about200 mL or less, e.g., about 150 mL or less, about 100 mL or less, about50 mL or less, about 30 mL or less, about 15 mL or less, about 10 mL orless, or about 5 mL or less. The formulation can be provided in a volumebounded by any two of the aforementioned endpoints. For example, theformulation can be provided in a volume of about 1 mL to about 200 mL,about 1 mL to about 50 mL, about 3 mL to about 30 mL, about 5 mL toabout 100 mL, or about 3 mL to about 10 mL. In certain preferredembodiments, the volume of the formulation is about 5 mL. One ofordinary skill in the art can readily select an appropriate containerbased upon the volume of the formulation.

The formulation comprises at least one stabilizing agent. Thestabilizing agent serves to stabilize levothyroxine or apharmaceutically acceptable salt thereof in the liquid formulation.

In some embodiments, the stabilizing agent is an amine. Non-limitingexamples of suitable amines include tromethamine (i.e.,2-amino-2-hydroxymethyl-propane-1,3-diol or Tris),bis(2-hydroxyethyl)-imino-tris(hydroxymethyl)methane (Bis-tris orBis-tris methane), monoethanolamine, diethanolamine, triethanolamine,2-amino-2-methyl-1,3-propanediol,2-dimethylamino-2-methyl-1-propanediol, 2-amino-2-ethylpropanol,2-amino-1-butanol, and 2-amino-2-methyl-1-propanol. Preferably, theamine is tromethamine.

The amine can be present in the formulation in any suitableconcentration. Typically, the amine can be present in the formulation ata concentration of about 1 mg/mL (milligram/milliliter) or more, forexample, about 5 mg/mL or more, about 10 mg/mL or more, about 15 mg/mLor more, or about 20 mg/mL or more. Alternatively, the amine can bepresent in the formulation at a concentration of about 50 mg/mL or less,for example, about 45 mg/mL or less, about 40 mg/mL or less, about 35mg/mL or less, about 30 mg/mL or less, about 25 mg/mL or less, or about20 mg/mL or less.

Thus, the amine can be present in the formulation in a concentrationbounded by any two of the aforementioned endpoints. For example, theamine can be present in the formulation in a concentration of about 1mg/mL to about 50 mg/mL, for example, about 1 mg/mL to about 50 mg/mL,about 5 mg/mL to about 45 mg/mL, about 5 mg/mL to about 40 mg/mL, about5 mg/mL to about 35 mg/mL, about 5 mg/mL to about 30 mg/mL, about 5mg/mL to about 25 mg/mL, or about 5 mg/mL to about 20 mg/mL. In apreferred embodiment, the amine is tromethamine present at aconcentration of about 20 mg/mL. In another preferred embodiment, theamine is tromethamine present at a concentration of about 10 mg/mL.

In some embodiments, the stabilizing agent is a salt of iodine, such assodium iodide or potassium iodide. In some embodiments, the formulationcomprises sodium iodide at a concentration of about 10 mcg/mL or more,e.g., 25 mcg/mL or more, 50 mcg/mL or more, 75 mcg/mL or more, 100mcg/mL or more, 125 mcg/mL or more, 150 mcg/mL or more, 175 mcg/mL ormore, or 200 mcg/mL or more. In other embodiments, the formulationcomprises sodium iodide at a concentration of about 500 mcg/mL or less,e.g., 450 mcg/mL or less, 400 mcg/mL or less, 350 mcg/mL or less, 300mcg/mL or less, 250 mcg/mL or less, 200 mcg/mL or less, 175 mcg/mL orless, or 150 mcg/mL or less.

Thus, the sodium iodide can be present in the formulation in aconcentration bounded by any two of the aforementioned endpoints. Forexample, the sodium iodide can be present in the formulation in aconcentration of about 10 mcg/mL to about 500 mcg/mL, for example, about50 mcg/mL to about 400 mcg/mL, about 100 mcg/mL to about 300 mcg/mL,about 125 mcg/mL to about 300 mcg/mL, about 125 mcg/mL to about 250mcg/mL, about 125 mcg/mL to about 200 mcg/mL, about 125 mcg/mL to about175 mcg/mL, or about 125 mcg/mL to about 150 mcg/mL. In a preferredembodiment, the sodium iodide is present at a concentration of about 140mcg/mL.

The formulation can comprise one, two, or three or more stabilizingagents. In certain embodiments, the formulation comprises an amine and asalt of iodine, preferably tromethamine and sodium iodide. In someembodiments, the formulation comprises about 10 mg/mL tromethamine andabout 140 mcg/mL sodium iodide.

The formulation comprises an isotonicity adjuster. Non-limiting examplesof suitable isotonicity adjusters include sodium chloride, potassiumchloride, dextrose, glycerin, and mannitol. In a preferred embodiment,the isotonicity adjuster is sodium chloride.

The isotonicity adjuster can be present at any suitable concentration.In some embodiments, the isotonicity adjuster is present at aconcentration that renders the formulation isotonic or approximatelyisotonic with cells (e.g., red blood cells) and/or isotonic orapproximately isotonic to blood plasma.

The formulation optionally comprises a pH adjuster. The pH adjuster canbe any suitable pH adjuster, for example, the pH adjuster can be sodiumhydroxide, potassium hydroxide, hydrochloric acid, or combinationsthereof. In a preferred embodiment, the pH adjuster is sodium hydroxide,hydrochloric acid, or a combination thereof.

The formulation can have any suitable pH. Typically, the formulation canhave a pH of about 9.0 or more including, for example, about 9.0 ormore, about 9.2 or more, about 9.4 or more, about 9.6 or more, about 9.8or more, about 10.0 or more, or about 10.2 or more. Alternatively, theformulation can have a pH of about 11.5 or less including, for example,about 11.3 or less, about 11.1 or less, about 11.0 or less, about 10.9or less, about 10.8 or less, about 10.7 or less, about 10.6 or less, orabout 10.5 or less.

The formulation can have a pH bounded by any two of the above endpointsrecited for the formulation. For example the formulation can have a pHof about 9.0 to about 11.5 including, for example, about 9.0 to about11.0, about 9.2 to about 10.8, about 9.2 to about 10.8, about 9.4 toabout 10.8, about 9.6 to about 10.8, about 9.8 to about 10.8, about 10.0to about 10.8, about 10.0 to about 10.7, about 10.0 to about 10.5, orabout 10.2 to about 10.6.

Tromethamine has a buffering range of about 7 to about 9. In a preferredembodiment, the pH of the formulation is about 9.8 to about 10.8, whichis above the buffering range of tromethamine. While not wishing to bebound by any particular theory, it is believed that tromethamine exertsa stabilizing effect on levothyroxine by a mechanism unrelated tobuffering of the formulation.

In a preferred embodiment, the formulation comprises (a) levothyroxineor a pharmaceutically acceptable salt thereof in a concentration ofabout 20 mcg/mL to about 100 mcg/mL, (b) tromethamine in a concentrationof about 5 mg/mL to about 20 mg/mL, (c) sodium iodide in a concentrationof about 100 mcg/mL to about 300 mcg/mL, (d) sodium chloride, and (e)water, wherein the formulation has a pH of about 9.8 to about 10.8.

The formulation that comprises levothyroxine or a pharmaceuticallyacceptable salt thereof, tromethamine, sodium iodide, sodium chloride,and water may further include one or more other substances. Non-limitingexamples of other substances include diluents, salts, buffers,stabilizers, solubilizers, and preservatives. In certain embodiments,the other substance is a cyclodextrin, such ashydroxypropyl-β-cyclodextrin or sulfobutylether β-cyclodextrin.

A formulation comprising levothyroxine or a pharmaceutically acceptablesalt thereof, tromethamine, sodium iodide, sodium chloride, and watercan be prepared by using any suitable technique, many of which are knownto those skilled in the art. The formulation can be prepared in a batchor continuous process. Generally, the formulation can be prepared bycombining the components thereof in any order. The term “component” asused herein includes individual ingredients (e.g., levothyroxine sodium,tromethamine, sodium iodide, sodium chloride, optional pH adjuster,etc.) as well as any combination of ingredients (e.g., levothyroxinesodium, tromethamine, sodium iodide, sodium chloride, optional pHadjuster, etc.). In some embodiments, the formulation is formed bycombining the components together in a vessel. The components can becombined in any order.

In some embodiments, the water is added to a suitable vessel, then thetromethamine, sodium iodide, and sodium chloride are added, eithersequentially or together, and the mixture is stirred. Next, the pH isadjusted to the desired value. Subsequently, the levothyroxine sodium isadded, and the mixture is stirred until the levothyroxine sodium isdissolved. In some embodiments, the water and sodium chloride arecombined and stirred until the sodium chloride is dissolved to providean aqueous solution of sodium chloride. Subsequently, the levothyroxinesodium, tromethamine, and sodium iodide are added, either sequentiallyor together, and the mixture is stirred. Next, the pH is adjusted to thedesired value. Optional ingredients, such as diluents, salts, buffers,stabilizers, solubilizers, and preservatives, can be provided to theformulation at any stage in its preparation.

In some embodiments, the formulation is filtered through one or morefilters prior to filling the composition into one or more suitablecontainers, such as a vial, an ampoule, a cartridge, a syringe, or abag. Preferably, one or more of the filtration steps and the fillingstep are performed under aseptic conditions in order to provide asterile container comprising a sterile formulation. A sterileformulation of the invention is preferably one in which substantiallyall forms of microbial life have been destroyed by an appreciable amountto meet the sterilization criteria set forth in the U.S. Pharmacopeia.See U.S. Pharmacopeia 32, NF 27, 1 (2009) 80-86.

The invention also provides a container comprising a formulationcomprising levothyroxine sodium, tromethamine, sodium iodide, sodiumchloride, optional pH adjustor, and any other optional components. Incertain embodiments, the container is a vial, an ampoule, a bag, abottle, a cartridge, or a syringe. In some embodiments, the container,the composition, or both the container and the composition are sterile.Preferably, the container is sealed by way of a closure, such as astopper, plunger, and/or tip-cap.

The container and closure can be made of glass, plastic, and/or rubber.One or more surfaces of the container and/or closure can be treated witha compound to limit reactivity with one or more components of theformulation. In some embodiments, the container and/or closure aretreated with silicon. In other embodiments, the container is treatedwith ammonium sulfate ((NH₄)₂SO₄). The container can be clear or opaque,and can be any color. In some embodiments, the container is flintcolored. In other embodiments, the container is amber colored.

In certain embodiments, the invention provides a pre-filled syringecontaining a formulation of the invention described herein. In certainembodiments, a syringe according to the invention is a component of anautoinjector.

In some embodiments, the liquid formulation of the invention containsnot more than 1.5% liothyronine (T3). In other embodiments, the liquidformulation contains not more than 1.25% liothyronine, e.g., not morethan 1.0% liothyronine, not more than 0.9% liothyronine, not more than0.8% liothyronine, not more than 0.7% liothyronine, not more than 0.6%liothyronine, not more than 0.5% liothyronine, not more than 0.4%liothyronine, not more than 0.35% liothyronine, not more than 0.30%liothyronine, not more than 0.25% liothyronine, not more than 0.2%liothyronine, or any range therein. For example, in certain embodiments,the liquid formulation contains 0.2%-1.5% liothyronine, 0.25%-1.25%liothyronine, 0.25%-1.0% liothyronine, 0.3%-0.9% liothyronine, 0.2%-0.4%liothyronine, 0.25%-0.4% liothyronine, or 0.25%-0.35% liothyronine.

In some embodiments, the liquid formulation contains not more than aspecified amount of liothyronine as measured after storage of theformulation at a predetermined temperature for a predetermined timeperiod. In certain embodiments, the liquid formulation contains not morethan 1.0% liothyronine, e.g., not more than 0.8% liothyronine, not morethan 0.6% liothyronine, not more than 0.5% liothyronine, not more than0.4% liothyronine, not more than 0.30% liothyronine, not more than 0.2%liothyronine, or any range therein as measured after storage of theformulation at 25±2° C. for a period of four months. In otherembodiments, the liquid formulation contains not more than 1.5%liothyronine, e.g., not more than 1.25%, not more than 1.0%, not morethan 0.8%, not more than 0.6%, not more than 0.5%, not more than 0.4%,or any range therein as measured after storage of the formulation at40±2° C. for a period of four months.

In some embodiments, the liquid formulation of the invention containsnot more than 5.0% total impurities. In other embodiments, the liquidformulation contains not more than 4.0% total impurities, e.g., not morethan 3.5% total impurities, not more than 3.0% total impurities, notmore than 2.5% total impurities, not more than 2.0% total impurities,not more than 1.5% total impurities, not more than 1.25% totalimpurities, not more than 1.0% total impurities, not more than 0.9%total impurities, not more than 0.8% total impurities, not more than0.7% total impurities, or any range therein. For example, in certainembodiments, the liquid formulation contains 1.0%-5.0% total impurities,1.5%-3.5% total impurities, 0.8%-3.0% total impurities, 0.7%-2.0% totalimpurities, 1.25%-4.0% total impurities, 0.8%-1.5% total impurities, or0.9%-1.25% total impurities.

In some embodiments, the liquid formulation contains not more than aspecified amount of total impurities as measured after storage of theformulation at a predetermined temperature for a predetermined timeperiod. In certain embodiments, the liquid formulation contains not morethan 2.0% total impurities, e.g., not more than 1.5% total impurities,not more than 1.25% total impurities, not more than 1.0% totalimpurities, not more than 0.9% total impurities, not more than 0.8%total impurities, not more than 0.7% total impurities, or any rangetherein as measured after storage of the formulation at 25±2° C. for aperiod of four months. In other embodiments, the liquid formulationcontains not more than 5.0% total impurities, e.g., not more than 4.0%total impurities, not more than 3.5% total impurities, not more than3.0% total impurities, not more than 2.5% total impurities, not morethan 2.0% total impurities, not more than 1.5% total impurities, or anyrange therein as measured after storage of the formulation at 40±2° C.for a period of four months.

The invention also provides a method of stabilizing a levothyroxineformulation by forming a mixture comprising levothyroxine or apharmaceutically acceptable salt thereof, tromethamine, sodium iodide,sodium chloride, and water, thereby stabilizing the formulation. Theidentity and amounts of levothyroxine or pharmaceutically acceptablesalt thereof, tromethamine, sodium iodide, and sodium chloride presentin the mixture as well as the pH can be the same as the identity andamounts of these components and the pH described herein with respect toa formulation of the invention. The formulation formed by the method ofstabilizing a levothyroxine formulation can have the same stabilitycharacteristics as the stability characteristics described herein withrespect to a formulation of the invention, particularly with regard tototal impurities and liothyronine.

The formulation according to the invention is suitable foradministration to a subject to treat or prevent a disease or condition.Preferably, the subject is a mammal. More preferably, the mammal is ahuman. Preferably, the disease or condition is a disease or conditionthat is treatable by the administration of levothyroxine or apharmaceutically acceptable salt thereof, such as hypothyroidism. Insome embodiments, the condition is myxedema coma.

The following examples further illustrate the invention but, of course,should not be construed as in any way limiting its scope.

Example 1

This example demonstrates the stability of exemplary formulationscomprising levothyroxine sodium, tromethamine, and water as a functionof the pH of the formulation.

Separate samples containing levothyroxine sodium at a concentration of20 mcg/mL, tromethamine at a concentration of 10 mg/mL in normal saline(0.9% NaCl in water) were adjusted to various pH levels. One sampleadditionally contained hydroxypropyl (HP) β-cyclodextrin at aconcentration of 10 mg/mL. 5 mL of each sample was filled into 10 ccamber tubing vials, and the vials were stoppered with 20 mm stoppersunder nitrogen. The samples were stored at temperatures of 25° C., 40°C. and 55° C. The samples stored at 55° C. were analyzed by HPLC at 1and 4 weeks (W) of storage. The samples stored at 40° C. were analyzedby HPLC at 4 W and 3 months (M) of storage. The samples stored at 25° C.were analyzed by HPLC at 3M of storage.

The HPLC conditions were as follows:

Column: Waters SYMMETRY™ C8 (5 μm, 4.6×150 mm) HPLC column

Mobile Phase A: Sodiumheptanesulfonate/Acetonitrile/Water/Methanol/H₃PO₄ (4.023 g/800 mL/1600mL/1600 mL/4 mL)

Mobile Phase B: Sodiumheptanesulfonate/Acetonitrile/Water/Methanol/H₃PO₄ (2.013 g/1000 mL/100mL/900 mL/2 mL)

Diluent: 0.01 N NaOH

Column temperature: 25° C.

Flow rate: 1.5 mL/min

Injection volume: 40-200 μL

Autosampler temperature: 5° C.

Detection: UV at 225 nm

Separation mode: Gradient

Gradient program:

Time % Mobile Phase (minutes) A B 25 100 0 40 10 90 50 10 90 51 100 0 60100 0The relative response time (RRT) for liothyronine to levothyroxine wasapproximately 0.73.

The results for liothyronine, largest unknown impurity, and totalimpurities as determined by peak area percent are set forth in Table 1.

TABLE 1 Levothyroxine Na (mcg/mL) 20 Tromethamine (mg/mL) 10HP-β-cyclodextrin (mg/mL) — — — — 10 Solvent Normal saline pH 8 9 9.5 109 55° C., 1 W % Liothyronine 15.2 2.7 1.2 0.6 2.7 % largest unknownimpurity 1.59 0.67 0.50 0.27 0.86 % total impurities 17.5 3.6 2.2 1.54.0 55° C., 4 W % Liothyronine nt nt 1.7 3.1 nt % largest unknownimpurity nt nt 0.09 0.55 nt % total impurities nt nt 2.1 4.2 nt 40° C.,4 W % Liothyronine nt nt 0.9 0.4 nt % largest unknown impurity nt nt0.12 0.12 nt % total impurities nt nt 1.7 0.6 nt 25° C., 3 M %Liothyronine nt nt 0.6 0.35 nt % largest unknown impurity nt nt 0.460.11 nt % total impurities nt nt 1.51 0.71 nt 40° C., 3 M % Liothyroninent nt 1.56 0.87 nt % largest unknown impurity nt nt 0.48 0.16 nt % totalimpurities nt nt 2.66 1.35 nt nt = not tested

The results described in Table 1 demonstrate reduced liothyronine andtotal impurities were detected in levothyroxine formulations having a pHof 9-10 as compared to pH 8.

The effect of pH on levothyroxine stability was further tested insamples having a pH 9.5-11.5. Separate samples containing levothyroxinesodium at a concentration of 20 mcg/mL or 100 mcg/mL, tromethamine at aconcentration of 10 mg/mL in normal saline were adjusted to various pHlevels. 5 mL of each sample was filled into 10 cc amber tubing vials,and the vials were stoppered with 20 mm stoppers under nitrogen. Thesamples were stored at temperatures of 25° C., 40° C., and 55° C. Thesamples stored at 55° C. were analyzed by HPLC at 1 W and 2 W ofstorage. The samples stored at 25° C. and 40° C. were analyzed by HPLCat 2M of storage using the HPLC conditions described hereinabove.

The results for liothyronine, largest unknown impurity, and totalimpurities as determined by peak area percent are set forth in Table 2.

TABLE 2 Levothyroxine Na (mcg/mL) 20 20 100 20 Tromethamine (mg/mL) 10Solvent Normal saline pH 9.5 10.4 10.4 11.5 55° C., 1 W % Liothyronine0.9 0.2 0.3 0.1 % largest unknown 0.09 0.09 0.11 0.08 impurity % totalimpurities 1.2 0.5 0.7 0.4 55° C., 2 W % Liothyronine 1.9 0.4 0.8 0.2 %largest unknown 0.1 0.1 0.11 0.1 impurity % total impurities 2.0 0.9 1.10.4 25° C., 2 M % Liothyronine 0.34 0.17 0.23 0.34 % largest unknown0.11 0.20 0.11 15.4 impurity % total impurities 0.7 0.6 0.6 19.6 40° C.,2 M % Liothyronine 1.00 0.38 0.49 0.31 % largest unknown 0.14 0.22 0.109.2 impurity % total impurities 1.44 0.88 0.8 12.1

The results described in Table 2 demonstrate that reduced liothyronineand/or total impurities were detected in levothyroxine formulationshaving a pH of 10.4 as compared to pH 9.5 or 11.5 following storage at25° C. or 40° C. for 2M.

Example 2

This example demonstrates the stability of exemplary formulationscomprising levothyroxine sodium, sodium iodide, and tromethamine as afunction of sodium iodide concentration and pH of the formulation.

Separate samples containing 20 mcg/mL levothyroxine sodium, 10 mg/mLtromethamine, 5.4 mg/mL sodium chloride, and sodium iodide at aconcentration of 280 mcg/mL, 140 mcg/mL, or 6 mcg/mL in water wereadjusted to various pH levels. 5 mL of each sample was filled into 10 ccflint molded vials, and the vials were stoppered with 20 mm stoppersunder nitrogen. The samples were stored at temperatures of 25° C. or 55°C. for 4 W prior to analysis by HPLC.

The HPLC conditions were as follows:

Column: ACE Excel 3 C18-PFP, 4.6×150 mm HPLC column

Mobile Phase A: Sodiumheptanesulfonate/Acetonitrile/Water/Methanol/H₃PO₄ (4.0 g/800 mL/1600mL/1600 mL/4.0 mL)

Mobile Phase B: Sodiumheptanesulfonate/Acetonitrile/Water/Methanol/H₃PO₄ (4.0 g/2000 mL/200mL/1800 mL/4.0 mL)

Diluent: 0.01 N NaOH

Column temperature: 25° C.

Flow rate: 1.5 mL/min

Injection volume: 80 μL

Autosampler temperature: 5° C.

Detection: UV at 225 nm

Separation mode: Gradient

Gradient program:

Time % Mobile Phase (minutes) A B 0 100 0 25 100 0 40 10 90 50 10 90 51100 0 60 100 0The relative response time (RRT) for liothyronine to levothyroxine wasapproximately 0.71.

The results for liothyronine, largest any other individual impurity(AOII), and total impurities as determined by peak area percent are setforth in Table 3.

TABLE 3 Levothyroxine Na (mcg/mL) 20 Tromethamine (mg/mL) 10 Solvent 5.4mg/mL sodium chloride in water NaI (mcg/mL) 280 140 6 pH 9.5 10 10.5 9.510 10.5 10.5 25° C., 4 W % Liothyronine 0.28 0.26 0.26 0.28 0.27 0.260.25 % AOII 0.56 0.55 0.57 0.52 0.55 0.6 0.33 % total impurities 1.581.61 1.66 1.51 1.64 1.75 1.9 55° C., 4 W % Liothyronine 1.63 1.06 0.711.74 1.06 0.74 0.87 % AOII 0.53 0.54 0.53 0.59 0.61 0.53 0.99 % totalimpurities 3.24 2.76 2.43 3.6 2.86 2.49 3.5

The samples also were stored at temperatures of 25° C. or 40° C. for 2Mor 4M prior to analysis by HPLC.

The HPLC conditions were as follows:

Column: Phenomenex Kinetex 2.6 μm C18, 4.6×150 mm HPLC column

Mobile Phase A: 0.05 M Sulfamic Acid, pH 2.0

Mobile Phase B: Acetonitrile

Diluent: 10% Mobile Phase A in Methanol:Acetonitrile:Mobile Phase A(1000 mL:300 mL:700 mL)

Column temperature: 27° C.

Flow rate: 1.2 mL/min

Injection volume: 50 μL

Autosampler temperature: 25° C.

Detection: UV at 225 nm

Separation mode: Gradient

Gradient program:

Time % Mobile Phase (minutes) A B 0 70 30 5 70 30 33 32 68 35 32 68 3670 30 50 70 30The relative response time (RRT) for liothyronine to levothyroxine wasapproximately 0.62.

The results for liothyronine, largest any other individual impurity(AOII), and total impurities as determined by peak area percent are setforth in Table 4.

TABLE 4 Levothyroxine Na (mcg/mL) 20 Tromethamine (mg/mL) 10 Solvent 5.4mg/mL sodium chloride in water NaI (mcg/mL) 280 140 6 pH 9.5 10 10.5 9.510 10.5 10.5 25° C., 2 M % Liothyronine 0.28 0.24 0.23 0.29 0.25 0.240.26 % AOII 0.11 0.12 0.11 0.09 0.10 0.12 0.46 % total impurities 0.810.8 0.85 0.76 0.77 0.88 1.85 40° C., 2 M % Liothyronine 0.9 0.55 0.390.88 0.55 0.41 0.43 % AOII 0.13 0.13 0.17 0.14 0.14 0.15 0.91 % totalimpurities 1.56 1.2 1.07 1.42 1.18 1.09 2.5 25° C., 4 M % Liothyronine0.35 0.28 0.25 0.35 0.28 0.26 0.25 % AOII 2.14 0.17 0.11 0.18 0.17 0.171.21 % total impurities 3.26 0.93 1.04 1.00 0.93 0.94 2.81 40° C., 4 M %Liothyronine 1.23 0.84 0.58 1.3 0.86 0.59 0.6 % AOII 0.52 1.38 1.07 0.720.92 0.69 1.45 % total impurities 2.76 3.23 2.72 2.84 2.73 2.14 3.51

The results described in Tables 3 and 4 demonstrate that levels ofliothyronine in formulations comprising 140 mcg/mL or 280 mcg/mL sodiumiodide were decreased as the pH was increased from 9.5 to 10 and from 10to 10.5 following storage at 55° C. for 4 W or at 40° C. for 2M or 4M.The levels of total impurities in formulations comprising 140 mcg/mLsodium iodide also were decreased as the pH was increased from 9.5 to 10and from 10 to 10.5 following storage at 55° C. for 4 W or at 40° C. for2M or 4M. Lower levels of AOII and total impurities were detected informulations comprising 140 mcg/mL or 280 mcg/mL sodium iodide at pH10.5 as compared to formulations comprising 6 mcg/mL sodium iodide at pH10.5 following storage at 55° C.° for 4 W or at 40° C. for 2M or 4M.

Example 4

This example demonstrates the stability of exemplary formulationscomprising levothyroxine sodium, sodium iodide, and tromethamine as afunction of vial type.

An aqueous solution containing 20 mcg/mL levothyroxine sodium, 10 mg/mLtromethamine, 5.4 mg/mL sodium chloride, and 6 mcg/mL sodium iodide wasadjusted to pH 10.5. 5 mL of the solution was filled into each of thevials described in Table 5, and the vials were stoppered under nitrogen.

TABLE 5 Inner Glass vial surface Type Size Color preparation treatmentVial 1 Glass 10 cc Flint Molded No Vial 2 Glass 10 cc Amber Molded(NH₄)₂SO₄ Vial 3 Glass  5 cc Amber Tubing (NH₄)₂SO₄ Vial 4 Glass  6 ccFlint Molded No Vial 5 Plastic¹ 10 cc Opaque N/A N/A Vial 6 Plastic² 10cc Opaque N/A N/A Vial 7 Plastic³ 10 cc Clear N/A N/A Vial 8 Plastic³ 10cc Amber N/A N/A Vial 9 Plastic⁴ 10 cc Clear N/A Silicon Vial 10Plastic⁴ 10 cc Amber N/A Silicon ¹polypropylene copolymer - ExxonMobilPP9122 ²polypropylene copolymer - Flint Hills Resources 23M2A ³cyclicolefin polymer - Daikyo CRYSTAL ZENITH ™ ⁴cyclic olefin polymer - SiO₂Medical Products

The vials were stored at a temperature of 25° C. or 55° C. for 4 W priorto analysis by HPLC. The HPLC conditions were the same as describedhereinabove for the data of Table 3. The results for liothyronine (T3),largest any other individual impurity (AOII), and total impurities asdetermined by peak area percent are set forth in Table 6.

TABLE 6 Storage Temp 25° C. 55° C. Impurity T3 AOII Tot T3 AOII Tot Vial1 0.25 0.33 1.9 0.87 0.99 3.5 Vial 2 0.28 0.39 2 1.59 4.97 14.6 Vial 30.25 2.54 4.4 0.91 2.31 4.9 Vial 4 0.22 0.52 2.5 1.11 4.02 7.9 Vial 50.25 0.7 3.4 0.85 8.76 20.3 Vial 6 0.25 0.89 3.7 0.73 10.51 29.9 Vial 70.24 0.75 2.6 0.78 7.29 10.7 Vial 8 0.24 0.81 2.7 0.75 7.09 10.6 Vial 90.25 0.48 1.9 0.78 2.79 5.4 Vial 10 NT NT NT 0.76 3 5.7

The results described in Table 6 demonstrate that vial material, size,color, and/or treatment can affect the stability of formulationscomprising levothyroxine sodium, sodium iodide, and tromethamine.

Example 5

This example demonstrates the stability of comparative formulationscomprising levothyroxine sodium, glycerol, sodium chloride, and water asa function of the pH of the formulation.

Separate samples containing levothyroxine sodium at a concentration of20 mcg/mL and glycerol at a concentration of 100 mg/mL in normal salinewere adjusted to pH levels of 7, 8, and 9. 5 mL of each sample wasfilled into 10 cc amber tubing vials, and the vials were stoppered with20 mm stoppers under nitrogen. The samples were stored at a temperatureof 55° C. The samples were analyzed by HPLC at 1 week of storage usingthe HPLC conditions described in Example 1. The results forliothyronine, largest unknown impurity, and total impurities asdetermined by peak area percent are set forth in Table 7.

TABLE 7 Levothyroxine Na (mcg/mL) 20 Glycerol (mg/mL) 100 Solvent Normalsaline pH 7 8 9 55° C., 1 W % Liothyronine 3.6 3.4 2.4 % largest unknown1.16 1.30 0.82 impurity % total impurities 5.3 5.4 3.8

The results described in Table 7 demonstrate that high levels ofimpurities are formed in levothyroxine formulations containing glycerolover the pH range 7-9 following storage at 55° C. for one week.

Example 6

This example demonstrates a method for preparing an exemplaryformulation of the invention.

The composition of an exemplary formulation containing 100 mcglevothyroxine in 5 mL volume is as described in Table 8.

TABLE 8 Component Quantity per mL Levothyroxine sodium, USP 20 mcgSodium chloride 6.48 mg Sodium iodide 0.14 mg Tromethamine, USP 10 mgSodium hydroxide (1N) As needed to adjust pH to 10-10.5 Hydrochloricacid (1N) (target 10.3) Purified water q.s.

The compositions for exemplary formulations containing 200 mcg or 500mcg levothyroxine in 5 mL volume are the same as described in Table 8,except that the concentrations of levothyroxine sodium are 40 mcg/mL and100 mcg/mL, respectively.

An exemplary formulation is prepared by filling purified water in anamount of approximately 80% of a predetermined final batch volume into asuitable container. The entire amounts of sodium chloride, sodiumiodide, and tromethamine are added in succession, with mixing untildissolution of each ingredient prior to addition of the next ingredient.The pH is determined, and then adjusted to pH 10.3 (range of 10.0 to10.5) with sodium hydroxide and/or hydrochloric acid. The entire amountof levothyroxine sodium is added to the container, and the solution ismixed until dissolution. The pH is determined, and then adjusted to pH10.3 (range of 10.0 to 10.5) with sodium hydroxide and/or hydrochloricacid. Purified water is added in an amount sufficient to reach thepredetermined batch volume with continued mixing to ensure completedissolution of all ingredients. The formulation can be bubbled withnitrogen or other suitable gas throughout the compounding to limit thedissolved oxygen in the formulation. Under aseptic conditions, thesolution is filtered through a 0.22 μm filter, and then 5 mL of thefiltered solution is filled into containers (e.g. vials) under nitrogen.The containers are then sealed (e.g., stoppered) under nitrogen.

All references, including publications, patent applications, andpatents, cited herein are hereby incorporated by reference to the sameextent as if each reference were individually and specifically indicatedto be incorporated by reference and were set forth in its entiretyherein.

The use of the terms “a” and “an” and “the” and “at least one” andsimilar referents in the context of describing the invention (especiallyin the context of the following claims) are to be construed to coverboth the singular and the plural, unless otherwise indicated herein orclearly contradicted by context. The use of the term “at least one”followed by a list of one or more items (for example, “at least one of Aand B”) is to be construed to mean one item selected from the listeditems (A or B) or any combination of two or more of the listed items (Aand B), unless otherwise indicated herein or clearly contradicted bycontext. The terms “comprising,” “having,” “including,” and “containing”are to be construed as open-ended terms (i.e., meaning “including, butnot limited to,”) unless otherwise noted. Recitation of ranges of valuesherein are merely intended to serve as a shorthand method of referringindividually to each separate value falling within the range, unlessotherwise indicated herein, and each separate value is incorporated intothe specification as if it were individually recited herein. All methodsdescribed herein can be performed in any suitable order unless otherwiseindicated herein or otherwise clearly contradicted by context. The useof any and all examples, or exemplary language (e.g., “such as”)provided herein, is intended merely to better illuminate the inventionand does not pose a limitation on the scope of the invention unlessotherwise claimed. No language in the specification should be construedas indicating any non-claimed element as essential to the practice ofthe invention.

Preferred embodiments of this invention are described herein, includingthe best mode known to the inventors for carrying out the invention.Variations of those preferred embodiments may become apparent to thoseof ordinary skill in the art upon reading the foregoing description. Theinventors expect skilled artisans to employ such variations asappropriate, and the inventors intend for the invention to be practicedotherwise than as specifically described herein. Accordingly, thisinvention includes all modifications and equivalents of the subjectmatter recited in the claims appended hereto as permitted by applicablelaw. Moreover, any combination of the above-described elements in allpossible variations thereof is encompassed by the invention unlessotherwise indicated herein or otherwise clearly contradicted by context.

The invention claimed is:
 1. A liquid formulation comprisinglevothyroxine or a pharmaceutically acceptable salt thereof; astabilizing agent comprising tromethamine; not more than 2% liothyronine(T3); and water; wherein the formulation retains at least about 95% ofthe initial concentration of levothyroxine or pharmaceuticallyacceptable salt thereof after storage for 12 months at 25±2° C., andretains at least about 95% of the initial concentration of levothyroxineor pharmaceutically acceptable salt thereof after storage for 2 monthsat 40±2° C.
 2. The formulation of claim 1, wherein levothyroxine or apharmaceutically acceptable salt thereof is levothyroxine sodium.
 3. Theformulation of claim 2, wherein levothyroxine sodium is present at aconcentration of from about 5 mcg/mL to about 500 mcg/mL.
 4. Theformulation of claim 1, wherein the tromethamine is present at aconcentration of about 1 mg/mL to about 50 mg/mL.
 5. The formulation ofclaim 1, wherein the stabilizing agent comprises a salt of iodine. 6.The formulation of claim 5, wherein the salt of iodine is sodium iodideor potassium iodide.
 7. The formulation of claim 6, wherein the salt ofiodine is sodium iodide which is present at a concentration of about 10mcg/mL to about 500 mcg/mL.
 8. The formulation of claim 1, wherein theformulation has a pH of from about 9.0 to about 11.5.
 9. The formulationof claim 8, wherein the formulation has a pH of from about 9.8 to about10.8.
 10. The formulation of claim 1, wherein the formulation containsnot more than 1.5% liothyronine (T3).
 11. The formulation of claim 1,wherein the formulation contains not more than 1.0% liothyronine (T3).12. The formulation of claim 1, wherein the formulation contains notmore than 5.0% total impurities.
 13. The formulation of claim 1, whereinthe formulation contains not more than 2.5% total impurities.
 14. Theformulation of claim 1, wherein the formulation retains at least about90% of the initial concentration of levothyroxine or pharmaceuticallyacceptable salt thereof after storage for at least 18 months at 25±2° C.15. The formulation of claim 1, wherein the formulation retains at leastabout 90% of the initial concentration of levothyroxine orpharmaceutically acceptable salt thereof after storage for at least 24months at 25±2° C.
 16. The formulation of claim 1, wherein theformulation does not contain a buffer.
 17. The formulation of claim 1,wherein the formulation is a ready-to-use formulation contained within avial, ampoule, cartridge, syringe, or bag.